The goal of the work proposed is to complete investigations which are necessary prior to testing gene therapy for acute lung injury in humans. The ultimate goal of clinical application is realistic because of rapid advancement of application of gene therapy in humans in other areas and because of the information obtained from the work completed during the current funding period of this grant demonstrating feasibility of the approach. We will use cationic liposomes to deliver DNA in the form of plasmids to the lungs either by intravenous injection or by aerosol administration. We will make new plasmid constructs containing either the Epstein-Barr eukaryotic replication initiation site or long terminal repeat sequences from adeno-associated virus as strategies to increase transgene expression and test the behavior of these vectors with reporter genes in cultured bovine pulmonary artery endothelial cells and in rabbits in vivo. We will use those data to construct vector which exuberantly express either the human mangano superoxide dismutase (MnSOD) or human alpha-1 antitrypsin (AAT) genes. We will determine the pathophysiologic effects of administration of plasmid/cationic liposome complexes either intravenously or by aerosol in rabbits and sheep. We will use plasmids containing the cDNA encoding either human MnSOD or human AAT in in vivo studies in rabbits and sheep to demonstrate and quantitate expression of these transgenes, to determine organ distribution of transgene expression and to determine cell sites of transgene expression. Finally, we will determine whether expressing either human MnSOD or human AAT in the lungs of rabbits and sheep will protect the lungs from the consequences of endotoxemia. Our hypothesis is that expression of transgenes encoding AAT or MnSOD can be accomplished in vivo and that this intervention will protect the lungs from injury by endotoxin. Based on the data collected so far, we also believe that this will prove a safe intervention which will provide a new category of therapeutic modalities in acute lung injury and other acute lung diseases.